Project Summary/Abstract Colon cancer is the 2nd-leading cause of cancer-related deaths in the United States and is one of the best- characterized solid tumors in terms of its common genetic mutations. However, knowledge gaps exist in the basic understanding of these cancers downstream of their genetic profile. For instance, the tumor-suppressor gene SMAD4 is mutated in approximately 57% of all colon tumors, and is associated with poor prognosis in patients, yet there is still little understanding of its molecular mechanisms in colon cancer. Additionally, while ~70% of colon cancers follow the typical WNT-driven adenoma-to-carcinoma pathway, approximately 20% follow the ?serrated tumor pathway?- often driven by gain-of-function BRAF mutations and have the worst prognosis of colon cancers. The proposal?s main objective will focus on elucidating the mechanistic role of the tumor suppressor SMAD4 in colon cancers. The hypothesis is that SMAD4 plays a critical transcriptional regulatory role in regulating the WNT pathway and is a key regulator in the serrated tumor pathway. The rationale is based on preliminary studies that reveal: 1) that SMAD4 binds to regulatory regions of the genome also bound by -catenin ? the transcriptional effector of the most commonly mutated signaling pathway in colon cancer, the WNT pathway; and 2) SMAD4 loss, when combined with activation of BRAF, can trigger serrated tumor formation as rapidly as 1 month in mice. Aim 1 will use epigenomic approaches to map the interaction of SMAD4 with -catenin at the level of DNA-binding - detailing the first intersection of these pathways on the colon cancer genome. Aim 2 will use new mouse models to determine how SMAD4 suppresses the serrated cancer transition from hyperplastic lesions to dysplasias. Aim 3 will use state-of-the- art tumor organoid model systems to determine how SMAD4 suppresses serrated cancer metastasis. The goal is to test the hypotheses that 1) SMAD4 directly regulates the WNT-signaling pathway by redirecting -catenin to tumor-suppressive gene targets in coordination with RUNX3, thus impacting cancer development, and 2) that SMAD4 suppresses key signaling pathways that are required for serrated tumor progression and metastasis. The proposed studies are significant in that they will identify the regulatory targets of a commonly mutated tumor-suppressor gene and present a new perspective on an understudied, but more deadly, colon cancer tumor type. These studies would have broad impacts in the cancer research field, and will reveal new targets to identify and treat patients with serrated tumors. With the co-mentorship of Drs. Michael Verzi and Ronald Hart, this proposal's training plan will prepare me for a transition to independence by 1) continuing to enhance my epigenomics skillset, 2) develop novel model systems to address critical questions in the colon cancer field, and 3) fortify the skills required to establish and maintain my own independent research program. The Department of Genetics at Rutgers University has been an outstanding institution for my postdoctoral training, and I will benefit greatly from the environment as I transition towards independence.